Immunophenotyping of Peripheral Eosinophils Demonstrates Activation in Eosinophilic Esophagitis

Abstract

Background and Aim Eosinophilic esophagitis (EoE) is a chronic inflammatory disorder characterized by upper gastrointestinal symptoms and the presence of high numbers of eosinophils in the esophagus. Although eosinophils in the esophagus have been found to be activated in subjects with EoE, detailed studies of intracellular signaling pathways involved in the mechanism of activation of eosinophils in EoE have heretofore been limited. The aim of the study was to assess whether any surface molecules or transcription factors are activated in peripheral eosinophils in subjects with EoE. Methods Eosinophils and CD3+ lymphocytes were identified directly from 50 μL of whole blood of EoE and control subjects. Using Hi-FACS, levels of surface activation markers, including CD66b, and intracellular phosphoepitopes, including phosphorylated forms of signal transducer and activator of transcription (phospho-STAT) 1 and 6, were measured within each cell subset. Results Levels of surface CD66b as well as levels of intracellular phospho-STAT1 and phospho-STAT6 in peripheral blood eosinophils were significantly higher for untreated subjects with EoE vs healthy controls (P <0.05). Levels of phospho-STAT1 and phospho-STAT6 in peripheral blood eosinophils were lower in subjects with EoE on therapy versus untreated subjects with EoE (P <0.05). Conclusions Levels of phospho-STAT1 and phospho-STAT6, transcription factors involved in inflammatory processes, were both significantly higher in peripheral eosinophils from untreated (ie, newly diagnosed) subjects with EoE versus subjects with EoE on therapy, healthy controls. Blood-based measurements of CD66b and phospho-STAT levels in peripheral eosinophils may be beneficial for identifying EoE.

Differences in eosinophil molecular profiles between children and adults with eosinophilic esophagitis

“…We have recently demonstrated that FOXP3 is mainly found in the cytoplasm of human eosinophils, akin to activated effector T cells, indicating that its expression may reflect eosinophil activation . Altogether, our data indicate that eosinophils are differentially activated in children and adults with EoE, and it is already known that eosinophils from children with EoE express the transcription factors STAT‐1 and STAT‐6 …”

“…18 Altogether, our data indicate that eosinophils are differentially activated in children and adults with EoE, and it is already known that eosinophils from children with EoE express the transcription factors STAT-1 and STAT-6. 28 The observation of distinct differences in molecular patterns between children and adults with EoE led us to investigate whether there might exist age-related differences in the levels of eosinophil molecules that were independent of disease. Indeed, we found that eosinophils from healthy individuals expressed distinctive molecular patterns depending on the age of the individual.…”

“…This “adult eosinophil EoE pattern” is characterized by increased levels of the surface markers CD11c, CD18, CD23, CD54, and CRTH2 and decreased levels of CD44 and CCR3 . A study of pediatric EoE revealed that the blood eosinophils from untreated patients had upregulated levels of surface CD66b and increased intracellular levels of the transcription factors “signal transducer and activator of transcription”‐1 (STAT‐1) and STAT‐6, both of which were phosphorylated . We have recently shown that eosinophils in the circulation of adult, untreated EoE patients have increased mRNA and protein levels of the transcription factor FOXP3 …”

“…26,27 A study of pediatric EoE revealed that the blood eosinophils from untreated patients had upregulated levels of surface CD66b and increased intracellular levels of the transcription factors "signal transducer and activator of transcription"-1 (STAT-1) and STAT-6, both of which were phosphorylated. 28 We have recently shown that eosinophils in the circulation of adult, untreated EoE patients have increased mRNA and protein levels of the transcription factor FOXP3. 18 In the present study, our first aim was to determine whether the blood eosinophils of children with EoE have a molecular pattern that is distinct from that of healthy children.…”

Non- and Semi-Invasive Methods of Monitoring Eosinophilic Esophagitis

“…Another study examined whether phenotypic analysis of eosinophil surface markers could distinguish treated from untreated disease. Elevated levels of surface CD66 intracellular phospho-STAT1, and phosphor-STAT6 were able to differentiate children with active disease from treated and healthy controls[24]. Finally, a prospective study of 47 children revealed that absolute eosinophil count, eosinophil derived neurotoxin and eotaxin-3 levels together correlated with esophageal inflammation[25].…”